Untargeted Assessment of Tumor Fractions in Plasma for Monitoring and Prognostication from Metastatic Breast Cancer Patients Undergoing Systemic Treatment

Abstract

The aim of this study was to assess the prognostic and predictive value of an untargeted assessment of tumor fractions in the plasma of metastatic breast cancer patients and to compare circulating tumor DNA (ctDNA) with circulating tumor cells (CTC) and conventional tumor markers. In metastatic breast cancer patients (n = 29), tumor fractions in plasma were assessed using the untargeted mFAST-SeqS method from 127 serial blood samples. Resulting z-scores for the ctDNA were compared to tumor fractions established with the recently published ichorCNA algorithm and associated with the clinical outcome. We observed a close correlation between mFAST-SeqS z-scores and ichorCNA ctDNA quantifications. Patients with mFAST-SeqS z-scores above three (34.5%) showed significantly worse overall survival (p = 0.014) and progression-free survival (p = 0.018) compared to patients with lower values. Elevated z-score values were clearly associated with radiologically proven progression. The baseline CTC count, carcinoembryonic antigen (CEA), and cancer antigen (CA)15-5 had no prognostic impact on the outcome of patients in the analyzed cohort. This proof of principle study demonstrates the prognostic impact of ctDNA levels detected with mFAST-SeqS as a very fast and cost-effective means to assess the ctDNA fraction without prior knowledge of the genetic landscape of the tumor. Furthermore, mFAST-SeqS-based ctDNA levels provided an early means of measuring treatment response.

Keywords: cell free circulating tumor DNA (ctDNA); circulating tumor cells (CTCs); mFAST-SeqS; metastatic breast cancer (MBC); prognosis; treatment response.

Figure 1

Correlation between mFast-SeqS based z-scores and ichorCNA tumor fractions. (A) With the exception of some samples (highlighted in red), the majority showed a high correlation between z-score and tumor fraction. Most of these outliers were obtained from patient B5. (B) Samples with high tumor fraction but lower z-scores revealed a lower number of copy number alterations (CNAs) compared to other samples as exemplified by two genome-wide copy number profiles. Based on ichorCNA, both samples had a tumor fraction of approximately 30%. However, due to the low abundance of CNAs in patient B5, the mFAST-SeqS z-score was only 8.4, compared to a value of 51.4 of a patient with many CNAs across the genome. Red indicates a gain of chromosomal material, blue indicates a loss of chromosomal material, green depicts balanced regions. (C) The removal of samples with low overall amounts of CNA significantly improved the correlation. (D) Correlation between mFAST-SeqS z-scores and ichorCNA-based tumor fractions in selected individual patients.

Figure 2

Kaplan–Meier survival analysis according to z-scores in metastatic breast cancer patients. Kaplan–Meier curves show that patients with z-scores ≥3 have significantly less (B) overall survival and (A) progression-free survival.

Figure 3

Longitudinal monitoring of z-scores and ichorCNA tumor fractions in circulating tumor DNA indicated for eight selected patients. Multiple measurements of tumor markers CEA and CA15-3 are also shown over time. Therapy regimens are indicated as colored shading and disease status at various times (ascertained on computed tomography or MRI) is shown as vertical dotted lines. SD denotes stable disease, PD progressive disease, PR partial response and MR minor response.

Figure 3

Longitudinal monitoring of z-scores and ichorCNA tumor fractions in circulating tumor DNA indicated for eight selected patients. Multiple measurements of tumor markers CEA and CA15-3 are also shown over time. Therapy regimens are indicated as colored shading and disease status at various times (ascertained on computed tomography or MRI) is shown as vertical dotted lines. SD denotes stable disease, PD progressive disease, PR partial response and MR minor response.